Triplet Imaging of Oxygen Consumption During the Contraction of a Single Smooth Muscle Cell (A7r5)

  • Matthias Geissbuehler
  • Thiemo Spielmann
  • Aurélie Formey
  • Iwan Märki
  • Marcel Leutenegger
  • Boris Hinz
  • Kai Johnsson
  • Dimitri Van De Ville
  • Theo Lasser
Conference paper
Part of the Advances in Experimental Medicine and Biology book series (volume 737)

Abstract

Triplet imaging is a novel optical technique that allows investigating oxygen metabolism at the single cell and the sub-cellular level. The method combines high temporal and spatial resolutions which are required for the monitoring of fast kinetics of oxygen concentration in living cells. Calibration and validation are demonstrated with a titration experiment using l-ascorbic acid with the enzyme ascorbase oxidase. The method was applied to a biological cell system, employing as reporter a cytosolic fusion protein of β-galactosidase with a SNAP-tag labeled with tetramethylrhodamine. Oxygen consumption in single smooth muscle cells A7r5 during an [Arg8]-vasopressin-induced contraction is measured. The triplet lifetime images over time can be related to an intracellular oxygen consumption corresponding to a mono-exponentially decaying intracellular oxygen concentration. This is in good agreement with previously reported measurements of oxygen consumption in skeletal muscle fibers.

References

  1. 1.
    Geissbuehler M et al (2010) Triplet imaging of oxygen consumption during the contraction of a single smooth muscle cell (A7r5). Biophys J 98(2):339–349PubMedCrossRefGoogle Scholar
  2. 2.
    Knopp JA, Longmuir IS (1972) Intracellular measurement of oxygen by quenching of fluorescence of pyrenebutyric acid. Biochim Biophys Acta – Gen Sub 279(2):393–397CrossRefGoogle Scholar
  3. 3.
    Vanderkooi JM et al (1987) An optical method for measurement of dioxygen concentration based upon quenching of phosphorescence. J Biol Chem 262(12):5476–5482PubMedGoogle Scholar
  4. 4.
    Hartmann P et al (1995) Luminescence quenching behavior of an oxygen sensor based on a Ru(II) complex dissolved in polystyrene. Anal Chem 67(1):88–93CrossRefGoogle Scholar
  5. 5.
    Mik EG et al (2006) Mitochondrial PO2 measured by delayed fluorescence of endogenous protoporphyrin IX. Nat Methods 3(11):939–945PubMedCrossRefGoogle Scholar
  6. 6.
    Sandén T et al (2007) Monitoring kinetics of highly environment sensitive states of fluorescent molecules by modulated excitation and time-averaged fluorescence intensity recording. Anal Chem 79(9):3330–3341PubMedCrossRefGoogle Scholar
  7. 7.
    Kautsky H, Müller G (1947) Luminescenzumwandlung durch Sauerstoff – Nachweis geringster Sauerstoffmengen. Z Naturforsch A 2(3):167–172Google Scholar
  8. 8.
    Rehm D, Weller A (1970) Kinetics of fluorescence quenching by electron and H-atom transfer. Isr J Chem 8(2):259–271Google Scholar
  9. 9.
    Lo LW (1996) Calibration of oxygen-dependent quenching of the phosphorescence of Pd-meso-tetra (4-carboxyphenyl) porphine: a phosphor with general application for measuring oxygen concentration in biological systems. Anal Biochem 236(1):153–160PubMedCrossRefGoogle Scholar
  10. 10.
    Renterghem CV (1988) Vasopressin modulates the spontaneous electrical activity in aortic cells (line A7r5) by acting on three different types of ionic channels. Proc Natl Acad Sci USA 85(23):9365–9369PubMedCrossRefGoogle Scholar
  11. 11.
    Keppler A et al (2003) A general method for the covalent labeling of fusion proteins with small molecules in vivo. Nat Biotechnol 21(1):86–89PubMedCrossRefGoogle Scholar
  12. 12.
    Hogan MC (2001) Fall in intracellular PO2 at the onset of contractions in Xenopus single skeletal muscle fibers. J Appl Physiol 90(5):1871–1876PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Matthias Geissbuehler
    • 1
  • Thiemo Spielmann
    • 1
    • 2
  • Aurélie Formey
    • 3
  • Iwan Märki
    • 1
  • Marcel Leutenegger
    • 1
    • 4
  • Boris Hinz
    • 5
  • Kai Johnsson
    • 6
  • Dimitri Van De Ville
    • 7
    • 8
  • Theo Lasser
    • 1
  1. 1.Laboratoire d’Optique Biomédicale LOBÉcole Polytechnique Fédérale de Lausanne (EPFL)LausanneSwitzerland
  2. 2.Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of TechnologyAlbaNova University CenterStockholmSweden
  3. 3.Laboratory of Cell Biophysics LCBÉcole Polytechnique Fédérale de Lausanne (EPFL)LausanneSwitzerland
  4. 4.Department of NanoBiophotonicsMax Planck Institute for Biophysical ChemistryGöttingenGermany
  5. 5.Laboratory of Tissue Repair and Regeneration, CIHR Group in Matrix DynamicsUniversity of TorontoTorontoCanada
  6. 6.Laboratory of Protein Engineering LIP1École Polytechnique Fédérale de Lausanne (EPFL)LausanneSwitzerland
  7. 7.Medical Image Processing Laboratory MIPÉcole Polytechnique Fédérale de Lausanne (EPFL)LausanneSwitzerland
  8. 8.Medical Image Processing Laboratory MIPUniversity of GenevaGenevaSwitzerland

Personalised recommendations